Tissue Kallikrein KLK1 Is Expressed de Novo in Endothelial Cells and Mediates Relaxation of Human Umbilical Veins

Abstract

AbstractBradykinin released by the endothelium is thought to play an important local role in cardiovascular regulation. However, the molecular identity of endothelial proteases liberating bradykinin from its precursors remained unclear. Using RTPCR and Southern blotting techniques we detected mRNA for tissue kallikrein (KLK1) in human umbilical vein endothelial cells and in bovine aortic endothelial cells. Protein expression was confirmed by precipitation of KLK1 from lysates of endothelial cells prelabeled with [35S]cysteine/methionine. Partial purification of tissue kallikrein from total endothelial cell extracts resulted in a protein triplet of about 50 kDa in Western blots using specific antiKLK1 antibodies. The immunodetection of tissue kallikrein antigen in the fractions from ion exchange chromatography correlated with the presence of amidolytic tissue kallikrein activity. Stimulation of endothelial cells with angiotensin II (ANGII), which recently has been shown to activate the vascular kinin system and to cause vasodilation, resulted in the release of bradykinin and kallidin. ANGIIdependent relaxation of preconstricted rings from human umbilical veins was abolished in the presence of a specific tissue kallikrein inhibitor. We conclude that endothelial cells de novo express significant amounts of tissue kallikrein, which likely serves in the local generation of vasoactive kinins.